Phosphorodithioic acid, mixed O,O-bis(1,3-dimethylbutyl and iso-Pr) esters, zinc salts

Short-term toxicity to fish

Administrative data

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

key study

Study period:

09 August 2004 - 10 March 2005

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Guideline study; restriction due to lack of analytical monitoring of test material

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2005

Report date:

2001

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method C.1 (Acute Toxicity for Fish)

Deviations:

no

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

Phosphorodithioic acid, mixed O,O-bis(1,3-dimethylbutyl and iso-Pr) esters, zinc salts

EC Number:

283-392-8

EC Name:

Phosphorodithioic acid, mixed O,O-bis(1,3-dimethylbutyl and iso-Pr) esters, zinc salts

Cas Number:

84605-29-8

Molecular formula:

Not applicable

IUPAC Name:

Phosphorodithioic acid, mixed O,O-bis(1,3-dimethylbutyl and iso-Pr) esters, zinc salts

Sampling and analysis

Analytical monitoring:

no

Details on sampling:

- Concentrations: 0, 1.0, 1.8, 3.2, 5.6 and 10 mg/L WAF loading rates

Test solutions

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Due to the low aqueous solubility and complex nature of the test material for the purposes of the definitive test the test material was prepared as a water accommodated fraction. Amounts of test material were each separately added via syringe to the surface of dechlorinated tap water to give the 1.0, 1.8, 3.2, 5.6 and 10 mg/L loading rates respectively. After the addition of the test material, the dechlorinated tap water was stirred by magnetic stirrer using a stirring rate such that a vortex was formed to give a slight dimple at the water surface. They were stirred for 24 hours. The stirring was stopped after 24 hours and the mixtures allowed to stand for 1 hour. The aqueous phase or WAF was removed by mid depth siphoning. Microscopic inspection of the WAFs showed no microdispersions or undissolved test material to be present.

- Controls: untreated negative control

- Evidence of undissolved material (e.g. precipitate, surface film, etc): No insoluble residues observed in the WAF test solutions.

Test organisms

Test organisms (species):

Oncorhynchus mykiss (previous name: Salmo gairdneri)

Details on test organisms:

TEST ORGANISM
- Common name: Rainbow trout
- Strain: Oncorhynchus mykiss
- Source: Fish were obtained from Brow Well Fisheries Limited, Hebden, Yorkshire UK and maintained in house since July 14 2004.
- Age at study initiation (mean and range, SD): juveniles
- Length at study initiation (length definition, mean, range and SD): mean standard length of 4.6 cm (sd=0.2)
- Weight at study initiation (mean and range, SD): mean weight of 1.23 g (sd=0.16) at the end of the definitive study
- Method of breeding: Fish were maintained in a glass fibre tank with a single pass water renewal system. Fish were acclimatized to test conditions from August 11 2004 to August 23 2004.

- Feeding during test
- Food type: commercial trout pellets
- Amount: no data available
- Frequency: Fish were fed commercial trout pellets until approximately 24 hours prior to the start of the definitive study

ACCLIMATION
- Acclimation period: Fish were acclimatized to test conditions from August 11 2004 to August 23 2004.
- Health during acclimation (any mortality observed): There was zero mortality in the 7 days prior to the start of the test.



QUARANTINE (wild caught)
- Duration: not applicable
- Health/mortality: not applicable

Study design

Test type:

semi-static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

96 h

Test conditions

Hardness:

100 mg/L as CaCO3

Test temperature:

14 degrees C (test temperatures ranged from 13.3 to 14.2

pH:

No treatment related differences in pH. pH values ranged from 7.4 to 8.1

Dissolved oxygen:

No treatment related differences in DO. Dissolved oxygen ranged from 9.4 to 10.2

Salinity:

not applicable

Nominal and measured concentrations:

Nominal concentrations of 0, 1.0, 1.8, 3.2, 5.6, and 10 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: 20 litre glass vessels
- Type (delete if not applicable): covered to reduce evaporation
- Material, size, headspace, fill volume:
- Aeration: no auxiliary aeration
- Type of flow-through (e.g. peristaltic or proportional diluter):
- Renewal rate of test solution (frequency/flow rate): semi-static, daily renewal of test preparations
- No. of organisms per vessel: 10
- No. of vessels per concentration (replicates): 1
- No. of vessels per control (replicates): 1
- Biomass loading rate: Test animals were equally and randomly assigned to the test vessels; The loading rate was determined to be 0.62 g/L

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Dechlorinated tap water passed through an activated carbon filter
- Metals: no data available
- Pesticides: no data available
- Chlorine: no data available
- Ca/mg ratio: Hardness: 100 mg/L CaCO3
- Culture medium different from test medium: No difference, the water in the culture system was the same type of water used in the bioassay. Fish were acclimated to test conditions prior to the study
- Intervals of water quality measurement: Daily for temperature, pH, DO

OTHER TEST CONDITIONS
- Adjustment of pH: none
- Photoperiod: 16 hours light and 8 hours dark with 20 minute dawn and dusk transition periods
- Light intensity: not reported

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) : Mortalities or sublethal effects of exposure were determined by visual inspection of the test fish after 3, 6, 24, 48, 72 and 96 hours

TEST CONCENTRATIONS
- Justification for using less concentrations than requested by guideline: NA

-Range finding study:
3 fish were added to each 20 litre test and control vessel and maintained at 14 degrees C.
- Test concentrations: 1, 10, and 100 mg/L nominal concentrations
- Results used to determine the conditions for the definitive study: The range finding results showed no mortalities at the 1.0 mg/L loading rate WAF. However mortalities were observed at the 10 and 100 mg/L loading rate WAFs. B ased on this information loading rates of 1.0, 1.8, 3.2, 5.6 and 10 mg/L.

Reference substance (positive control):

not specified

Results and discussion

Effect concentrations

Effect concentrations 1

Duration:

96 h

Dose descriptor:

LL50

Effect conc.:

4.5 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Remarks on result:

other: 3.5 - 5.7 mg/L

Effect concentrations 2

Duration:

96 h

Dose descriptor:

NOELR

Effect conc.:

1.8 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Details on results:

- Behavioural abnormalities: Sublethal effects of exposure were observed at the 5.6 mg/L loading rate WAF and above. These responses were increased pigmentation and moribund fish.

- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: No insoluble materials were observed in the test solutions. Microscopic inspection of the WAFs showed no microdispersions or undissolved test material to be present.

Results with reference substance (positive control):

No reference tox. data reported

Reported statistics and error estimates:

An estimate of the LL50 values at 3 and 6 hours was given by inspection of the mortality data. The LL50 values at 24 hour and the LL50 values and associated confidence limits at 48, 72 and 96 hours were calculated by the maximum likelihood probit method using the ToxCalc computer software package.

Any other information on results incl. tables

Sublethal observations / clinical signs:

Table1. Cumulative Mortality Data

Nominal Test Concentration mg/L	Cumulative Mortality at Hour (Initial population =10 fish)    3      6     24    48   72   96						% Mortality at 96 hours
0, Control	0	0	0	0	0	0	0
1.0	0	0	0	0	0	0	0
1.8	0	0	0	0	0	0	0
3.2	0	0	0	2	2	2	20
5.6	0	0	2	7	7	7	70
10	0	0	4	10	10	10	100

 

 

Table 2. Statistical Analysis of Mortality Data

Exposure Period (hours)	LL50 (mg/L)	95% Confidence Limits (mg/L)
3	>10	--
6	>10	--
24	10	--
48	4.5	3.5 to 5.7
72	4.5	3.5 to 5.7
96	4.5	3.5 to 5.7

Applicant's summary and conclusion

Conclusions:

The acute toxicity of the test material CAS# 84605-29-8 to the freshwater fish, Oncorhynchus mykiss, was investigated and resulted in a 96 hour LL50 of 4.5 mg/L and a NOEL of 1.8 mg/L.

Executive summary:

Introduction. A study was performed to assess the acute toxicity of the test material CAS# 84605-29-8 to rainbow trout using OECD 203 guidelines.

 

Methods. Following preliminary range finding studies fish were exposed in groups of ten to water accommodated fractions of the test material over a range of nominal loading rates of 1.0, 1.8, 3.2, 5.6, and 10 mg/L for a period of 96 hours at a temperature of 14 degrees C under semi static test conditions. The number of mortalities and any sublethal effects of exposure in each test and control vessel were determined 3 and 6 hours after the start of exposure and then daily throughout the test until termination after 96 hours.

 

Results.  The acute toxicity of the test material to the freshwater fish,Oncorhynchus mykiss, was investigated and resulted in a 96 hour LL50 of 4.5 mg/L (95% confidence limits of 3.5 to 5.7 mg/L) and a NOEL of 1.8 mg/L.